Indications of prostate cancer are predominantly an elevated prostate specific antigen (PSA) test or abnormal digital rectal exam (DRE), and prompt transrectal ultrasound (TRUS) guided biopsy of the prostate for definitive diagnosis. Often, there are cancers that are not visible (iso- echoic) on TRUS. Furthermore, in PSA screened populations, the accuracy of TRUS is only 52% due to false-positive results and is not reliable for the detection and localization of prostate cancer which is currently the only choice to perform biopsies. The aim is to exclude prostate cancer in patients with false positive PSA results and localize the cancer for targeted biopsies. To differentiate cancerous from benign prostate tissue, we propose to develop a handheld transrectal imaging probe based on multispectral photoacoustic (PA) imaging that can detect changes in relative concentrations of deoxyhemoglobin (dHb), oxyhemoglobin (HbO2), lipid and water in vivo. Specific Aims of our project include: Aim 1: To develop a dual modality system that will incorporate ultrasound (US) imaging into our existing ex-vivo PA imaging probe to generate co-registered dual modality images. Aim 2: To design and fabricate an in-vivo transrectal dual mode (US + PA) imaging probe capable of generating co-registered images suitable for prostate biopsy. Aim 3: Perform PA and US imaging experiments on the tissue equivalent phantoms to validate the performance and determine the quality metrics of the in-vivo dual mode imaging probe. The ultimate goal of this project is to develop an imaging modality that can clearly differentiate cancerous from benign prostate tissue in patients with positive PSA results. This will also help identify and monitor patients who need no further treatment, as well as aid in reducing false negative results due to poor cancer targeting of TRUS biopsy needles. This probe may help to reduce the TRUS biopsy rate or eliminate it altogether (if aggressive versus indolent cancers can be identified by multispectral photoacoustic imaging). Due to the functional nature of PA imaging, it is possible that the proposed probe may be a useful tool to evaluate prostate tumor prognosis, response to drugs, and detection of tumor recurrence.